Gut, 1976, 17, 561-567

Tissue demonstration of carcinoembryonic antigen (CEA) in ulcerative colitis P. ISAACSON From the Department of Pathology, Southampton University Hospital, Southampton

An immunoperoxidase technique was used to demonstrate carcinoembryonic antigen (CEA) in sections of proctocolectomy specimens from nine cases of ulcerative colitis in some of which carcinoma and premalignant change had developed. Carcinoma and premalignant change stained positively for CEA in contrast with benign mucosa, whether or not inflammatory dysplasia was pre-

SUMMARY

sent.

Using an immunoperoxidase technique to demonstrate carcinoembryonic antigen (CEA) in formalin fixed, paraffin embedded tissue, Isaacson and Le Vann (1976) have shown that carcinoma can be reliably differentiated from benign mucosa even when

the latter showsconsiderableinflammatorydysplasia. With the use of this technique it was also shown that, in adenomatous polyps, mucosa considered to be premalignant could be discriminated from benign adenomatous glands. In either case, malignant or premalignant mucosa stained strongly for CEA, while benign glands stained either very weakly or not at all. In this study cases of ulcerative colitis were investigated by this technique to determine whether foci of premalignant change could be identified by an increased content of CEA in comparison with surrounding inflamed mucosa. Methods

Proctocolectomy specimens from nine cases of ulcerative colitis were examined and their characteristics are summarized in the Table. All the specimens were received fresh from the operating theatre and immediately fixed in 10 % buffered formol saline. An average of 20 sections was examined from each specimen to include, where appropriate, multiple examples of carcinoma, precancerous change, and uncomplicated chronic ulcerative colitis. In deciding whether precancer was present the criteria used were those elaborated by Morson and Pang (1967) and Yardley and Keren (1974). Premalignant change was identified macroscopically as slightly raised plaquelike patches on the mucosa or as sessile villous adeReceived for publication 12 March 1976

561

Table Details of cases of ulcerative colitis Case

Age

no.

(yr)

1 2 3

63 54 33

4 5 6 7 8 9

Sex

Carci-

Pre-

noma

cancer

F M M

+ + +

+ + +

28

F

+

-

55 38 34 42 67

F M M F F

+

+ -

-

Other features

Multiple carcinomas Multiple carcinomas Multiple carcinomas Diffuse premalignant change Ileorectal anastamoses -

nomatous structures. Microscopically, there was usually a villous pattern (Fig. 2) and epithelial atypia in the form of goblet cell depletion, with hyperchromatic, often stratified, nuclei containing prominent nucleoli. No attempt was made to distinguish between premalignant change and carcinoma-in-situ, although in some instances the cytological atypia was severe enough to suggest malignancy (Fig. 3a). Therewasusuallylittle accompanying inflammation. The dysplastic changes induced by acute or chronic inflammation (Figs. 5a, 6a) did not give rise to a villous pattern. The glands were more regular but goblet cell depletion was present. The nuclei contained prominent nucleoli but were only moderately hyperchromatic and showed little stratification. There was often a greatly increased number of mitotic figures (Fig. 6a). Goat antihuman CEA and purified CEA were obtained from the Chester Beatty Research Institute, London, through the generosity of Professor A. Munro Neville. Details of the preparation of the

P. Isaacson

562

CEA and antiserum have been previously described (Isaacson and Le Vann, 1976). Sections were stained for CEA by an indirect immunoperoxidase technique using goat antiCEA followed by peroxidase conjugated horse antigoat immunoglobulin prepared by the method of Nakane and Kawaoi (1974). Serial dilutions of goat antiCEA serum were used to stain a control slide of colonic carcinoma and the dilution one step below that at which the reaction disappeared was used for the study. This dilution varied with each batch of peroxidase conjugated horse antigoat serum but was most often 1 in 200. Slides were pretreated with undiluted normal horse serum to prevent nonspecific staining and were counterstained with haematoxylin. The details of method used were otherwise as previously described (Isaacson and Le Vann, 1976). The same case of carcinoma with adjacent normal mucosa was used as a positive control throughout. The characteristic staining of CEA in colonic tissue

had previously been shown to be blocked by absorption of the goat antiCEA with purified CEA, and in the study normal goat serum was substituted for goat antiCEA as a negative control. Results Where CEA could be demonstrated in the colonic mucosa it appeared as a thin, slightly irregular line around glandular lumens or over the surface epithelium (Fig. 1). In an occasional cell the cytoplasm showed pale brown staining. Only carcinoma and mucosa showing premalignant change stained positively for CEA. Areas of active or chronic inactive ulcerative colitis were negative even when considerable inflammatory dysplasia was present (Figs. 2, 3, 4, 5, 6). Where Paneth cell metaplasia occurred these cells stained positively (Fig. 4), as they did in the section of ileum from the ileorectal anastamosis in case 4.

AV

(a)

(b)

Fig. 1 Case 4. (a) Rectal carcinoma. H and E, x 100. (b) Same field stained for CEA which shows up as a slightly irregular dark brown lumenal line. x 100.

(b) (a) Fig. 2 Case 3. (a) Mucosa showing premalignant change. Note villous pattern and cytological atypia. H and E, x 100. (b) Same field stained for CEA with positive result. x 100.

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i4

x6 ,

(b) (a) Fig. 3 Case 3. (a) High power view of a gland, part of which shows premalignant change with considerable atypia. H and E, x 400. (b) Same gland stained for CEA which is confined to the portion of the gland showing premalignant change. x 400.

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